The objective of the proposed research is to study the effect of acidic stress on cyanobacteria. The long term objectives of this laboratory have been to relate cyanobacterial growth and physiology to environmental variables and to determine the mechanisms by which cyanobacteria regulate the expression of their genes under stressful conditions. The hypotheses to be tested in the proposed research are that cyanobacteria have reproducible and specific responses to acidic stress, that they display an acid tolerance response to changes in external pH that allows the regulation of homeostasis, and that specific proteins are induced in response to acidic stress. 31P NMR spectroscopy will be used to determine internal pH of cells in various medium pHs and cell densities. 23Na NMR spectroscopy, using aqueous shift reagents, will allow the measurement of intracellular sodium concentrations in order to determine if a cation/proton antiporter is involved. Viability will be assessed using fluorescent microscopy and image analysis. One- and two-dimensional gel electrophoresis and autoradiography will be carried out to determine the various proteins that are induced during acid tolerance and stress. Granules that appear when cells are acid stressed will be studied and identified by confocal microscopy, NMR spectroscopy and chemical assay. How bacteria survive in stressful environments is an intriguing biological problem that will lead to understanding the basic cellular biology and physiology of both pathogenic and environmentally important microorganisms. New functions for cellular proteins may be described from the proteomic studies.